The anthrax letter attacks after September 11, 2001 demonstrated the ease of using the anthrax spore as a weapon in a bioterrorist attack. Anthrax is a lethal infectious disease caused by the spore-forming Bacillus anthracis. The two major virulence factors of B. anthracis are the poly-?-D-glutamic acid (?-D-PGA) capsule and the exotoxin. The three components of the exotoxin, protective antigen (PA), lethal factor (LF) and edema factor (EF) are secreted separately but form two bipartite toxins that cause massive edema and organ failure. The antiphagocytic ?-D-PGA capsule disguises the bacilli from immune surveillance by macrophages and allows unimpeded growth of the bacilli in the host; leading to anthrax disease caused by both the capsule and toxins. The currently licensed Anthrax Vaccine Adsorbed (AVA) or rPA based anthrax vaccines only address the toxin-induced disease, not the capsule mediated virulence. An antibody response triggered specifically by ?-D-PGA can fully neutralize the disguising capsule. However, the capsule is not immunogenic by itself and must to be conjugated to a carrier protein such as PA. Conjugation of ?-D-PGA to PA can provide immunogenic protection against both capsule and toxins. Based on successful efforts to experimentally demonstrate the anti-capsule properties generated by conjugated ?-D-PGA vaccine, we propose to undertake advanced development of two conjugate vaccines. A PGA-rPA conjugate vaccine and also the development of a conjugate vaccine targeting the receptor binding site of PA, Domain IV (PAD4), which due to its exposed location may be critical for inhibiting infection. PA has been a target of anthrax vaccine development because it can elicit a toxin-specific protective immune response that neutralizes PA and LF. However, efforts have been made to include ?-D-PGA as a component of 2nd generation anthrax vaccines, resulting in a promising vaccine candidate that can provide comprehensive protection. In this proposal, we will focus on the advanced development of such a promising countermeasure containing the two major virulence factors of B. anthracis (?-D-PGA and PA or PAD4) to deliver a safe and efficacious anthrax countermeasure. The conjugated vaccine concept embodies the paradigm of combining both antibacterial (prophylactic) and antitoxic (therapeutic) components into a single vaccine. There has been much success in inducing protective levels of antibodies in infants and children against systemic infection with encapsulated pathogens, and so rapid development of this conjugated anthrax vaccine will fill an urgent void, delivering a well-defined and characterized biodefense countermeasure potentially suitable for infants and children. The proposed work will lead to the characterization, testing and manufacturing of a promising anthrax vaccine candidate; creating the basis for subsequent advanced toxicology studies, submission of an Investigational New Drug application and safety testing in human clinical trials.